Prion protein (PrP) underlies a spectrum of diseases with no established treatment and devastating human and economic consequences. While substantial progress has been made in understanding the biochemical nature of the prion infectious agent, the lack of detailed structural information about the aggregated, disease-related forms of the prion protein (PrPSc) is a major barrier to progress in the field. In the current application, we propose to elucidate the structures of ordered, disease-related prion protein aggregates using state-of-the-art techniques including solid-state NMR methods supplemented with mass-per-length and volume-per-length measurements. The three specific aims are focused on elucidating the molecular structures of three homogeneous but conformationally distinct forms of the ordered PrP aggregates prepared in vitro using highly pure full-length recombinant PrP under well-defined conditions. They include PrPSc with a high infectivity titer and two types of PrP amyloid fibrils with limited infectivity or no detectible infectivity, respectively. When the goals of this project are accomplished, the current study will identify structural features that distinguish infectious and non- infectious states of self-propagating ordered PrP aggregates and define molecular determinants of prion infectivity. Ultimately, insight gained from this research will aid in the efforts to develop effective therapeutics for prion diseases and a better understanding of the mechanisms of prion replication. Furthermore, these studies will lay the ground work for understanding the mechanisms responsible for prion-like replication of abnormal conformation of non-prion proteins associated with other neurodegenerative diseases. PUBLIC HEALTH RELEVANCE: Prion diseases are a group of fatal age-dependent neurodegenerative diseases that can arise spontaneously or be inherited, but can also be infectious. Lack of structural information about infectious, disease-related forms of the prion protein represents the major critical barrier to progress in the field. The current project seeks t elucidate the molecular structure of disease-related forms of the prion protein and define molecular determinants of prion infectivity.